It is well known that part of the immune reaction in man and other animals is provided by antibodies: these are proteins which form part of the immunoglobulins produced by the lymphoid cells and which circulate in the blood. Antibodies combine chemically with antigens so as to neutralize them; however, it has been realized that, in many conditions, the antibodies do not provide the necessary degree of immunity and increasingly the importance is being recognized of the part played by the lymphocytes in providing cellular immunity. The lymphocytes also carry the immunological memory, and they provide a natural defence against viral infections and against the growth of neoplasms. Although some medical conditions are characterized by a deficiency of cellular immunity, treatment by transfusion of lymphocytes is unsatisfactory for two reasons: (a) a suitable donor is needed; and (b) the transfused lymphocytes can produce a transplant reaction in the host.
The presence in lymphocytes of a substance now commonly referred to as "transfer factor," and the role of this substance as carrier of the cellular immune memory, was first demonstrated by H. S. Lawrence [Lawrence H. S., "The transfer in humans of delayed skin sensitivity to streptococcal M substance and to tuberculin with disrupted leucocytes" J. Clin. Invest (1955) 34, 219-30; Lawrence H. S., Al Askari S., and David J., "Transfer of immunological information in humans with dialysates of leucocyte extracts," Trans. As. Amer. Physicians (1963) 76, 84-91; Lawrence H. S., "Transfer factor and cellular immune deficiency diseases," New Engl. J. Med. (1970) 283, 411-9; and Lawrence H. S., "Transfer factor" Advances in Immunology (1969), 11, 195-266]. It has since been demonstrated that the substance referred to as "transfer factor" is, in fact, a mixture of substances (which may be defined by chemical properties, spectrophotometric profile and, mainly, the biological property of transferring cellular immunity against one or a plurality of antigens) and is, therefore, more properly referred to as "transfer factors." This substance, which has been found in man and other primates, can transfer only cellular immune responses, and not the ability to secrete anitibodies. After contact with an antigen, the lymphocyte memory will contain transfer factors, which are currently believed to be specific to the class of antigens to which belongs the antigen with which the lymphocyte was contacted. Thus, transfer factors are specific to a class of antigens and exclusively to cellular immunity. However, the lymphocytes of a normal adult will contain transfer factors corresponding to each of the viral, bacterial or other antigens with which he has been in contact, so that dialysates of leucocyte extracts will contain a polyspecific transfer factor mixture.
Dialysis of transfer factors will divide them into dialysable transfer factor and non-dialysable transfer factor. Available evidence shows that dialysable transfer factor appears to be a small molecule, whose approximate molecular weight is estimated to be in the range of about 7,000; in fact, dialysable transfer factor itself would appear to be a mixture of molecules having various molecular weights, the majority being close to about 7,000 and the numbers of molecules with molecular weights substantially below about 4,000 or substantially above about 10,000 being negligible. Dialysable transfer factor appears to contain a ribonucleotide, but is not destroyed by ribonuclease. It has been shown to be stable for indefinite periods when lyophilized and kept at -20.degree. C. On the hand, nondialysable transfer factor is a larger molecule and has been characterized as RNA having a molecular weight greater than 30,000; both are not species-specific but are specific to an antigen class and are also specific to the transfer of cellular immunity.
Since dialysable transfer factor is not antigenic, that is to say that it cannot induce an immune reaction, its therapeutic administration would be preferable to the transfusion of lymphocytes. Indeed, experimental treatment with dialysable transfer factor has already proven successful in remedying deficiencies of cellular immunity and in obtaining amelioration of various diseases; thus, it has been used with indications of success in the treatment of subacute sclerosing panencephalitis, systemic candidiasis, malignant melanoma, breast cancer, measles and chickenpox, although it will be appreciated that this is by no means an exhaustive list of the diseases against which such treatment is effective. However, the prior art methods of producing transfer factors, which rely on donor blood as the starting material, do not permit transfer factors to be produced satisfactory for use as therapeutic agents: the production of any pure substance from whole blood is a tedious operation, requiring the preliminary separation of the lymphoid cells from a large volume of selected human blood.
It has been proposed to culture lymphocyte cells in vitro [Lawrence H. S., "Transfer factor" Advances in Immunology (1969) 11, 195-266] but this is primarily for the purpose of maintaining the lymphocytes alive and the transfer factor in usable condition after the lymphocyte cells have been removed from the donor and also to enable experiments to be carried out on those cells which could not conveniently or ethically be carried out in vivo in humans. Also, Jachertz in British Patent No. 1,229,888 discloses that a material he terms "informationally active RNA" can be synthesized in vitro and the product may be used as a "vaccine." The "informationally active RNA" of Jachertz is said to be produced by various "immunologically competent cells," including human and monkey leukocytes; Jachertz discloses that the immunologically competent cells may be cultured before the RNA is extracted therefrom. However, the cell culture is only carried out for a short time (5-60 minutes) and effectively does no more than maintain the cells alive: the process could not form the basis for commercial large-scale production of transfer factor. Furthermore the non-dialysable "informationally active RNA" is not the same product as transfer factor.
Thus, none of the prior art methods lends itself to the large-scale production of transfer factor under standardized, reproducible conditions. Moreover, because they use donor blood, these methods can only produce transfer factors which are specific to those classes of antigen with which the donor has been in contact; they cannot be modified to produce a particular antigen-specific transfer factor at will, except by choice of an appropriate blood donor.
Lymphocytes and other related cells cannot be maintained in cell culture for substantial periods of time without modification. It is, however, well-known that such cells may be transformed in such a way that they can be maintained in cell culture indefinitely. However, in the course of this modification, substantial changes in the character and properties of the cells are observed; the resulting cells are termed "lymphoblastoid cells." Although derived from lymphocytes, they share very few of the properties of native lymphocytes and are regarded by those skilled in the art as a totally different type of cell. We have now discovered that cultures of blastoides can be artificially induced to produce transfer factor, even if transfer factor was not previously present in the cells.
It is, therefore, one of the principal objects of Applicants invention to provide a process for producing transfer factor which does not rely upon donor blood and which can be used for the production at will of large quantities of any particular antigen-specific transfer factors desired.
It is a further and more specific object of the invention to provide a process in which lymphoblastoid cells are induced to produce transfer factor.